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AssigneeLife Technologies Corporation
InventorsChieh-Yuan Li, David Ruff, Shiaw-Min Chen, Jennifer O'Neil, Rachel Kasinskas, Jonathan Rothberg, Bin Li, Kai Qin Lao, Wolfgang Hinz
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In some embodiments, the present teachings provide methods for nucleic acid amplification, comprising forming a reaction mixture, and subjecting the reaction mixture to conditions suitable for nucleic acid amplification. In some embodiments, methods for nucleic acid amplification include subjecting the nucleic acid to be amplified to partially denaturing conditions. In some embodiments, methods for nucleic acid amplification include amplifying without fully denaturing the nucleic acid that is amplified. In some embodiments, the methods for nucleic acid amplification employ an enzyme that catalyzes homologous recombination and a polymerase. In some embodiments, methods for nucleic acid amplification can be conducted in a single reaction vessel. In some embodiments, methods for nucleic acid amplification can be conducted in a single continuous liquid phase of a reaction mixture, without need for compartmentalization of the reaction mixture or immobilization of reaction components. In some embodiments, methods for nucleic acid amplification comprise a amplifying at least one polynucleotide onto a surface under isothermal amplification conditions, optionally in the presence of a polymer. The polymer can include a sieving agent and/or a diffusion-reducing agent.
A composition for nucleic acid amplification comprising a recombinase, a recombinase accessory protein, a plurality of supports having a plurality of forward primers attached thereto, a plurality of reverse primers in solution, and a T5 or T7 DNA polymerase, wherein the plurality of forward primers have identical nucleotide sequences, wherein the plurality of reverse primers have identical nucleotide sequences, wherein the composition is a single continuous liquid phase comprising a sieving agent, wherein the single continuous liquid phase is not within a gel or matrix, wherein the T5 or T7 DNA polymerase has reduced exonuclease activity compared to wild-type T5 or T7 polymerase, respectively, and wherein if the polymerase is T7 polymerase, the composition further comprises thioredoxin.
A kit for nucleic acid amplification, comprising:
Dependencies are inferred from the claim text.